Membrane targeting properties of a herpesvirus tegument protein-retrovirusGag chimera

The retroviral Gag protein is capable of directing the production and relea se of virus-like particles in the absence of all other viral components. Bu dding normally occurs after Gag is transported to the plasma membrane by it s membrane-targeting and -binding (M) domain. In the Rous sarcoma virus (RS V) Gag protein, the M domain is contained within the first 86 amino acids. When M is deleted, membrane association and budding fail to occur. Budding is restored when M is replaced with foreign membrane-binding sequences, suc h as that of the Src oncoprotein. Moreover, the RSV M domain is capable of targeting heterologous proteins to the plasma membrane. Although the soluti on structure of the RSV M domain has been determined, the mechanism by whic h M specifically targets Gag to the plasma membrane rather than to one or m ore of the large number of internal membrane surfaces (e.g., the Golgi appa ratus, endoplasmic reticulum, and nuclear, mitochondrial, or lysosomal memb ranes) is unknown. To further investigate the requirements for targeting pr oteins to discrete cellular locations, we have replaced the M domain of RSV with the product of the unique long region 11 (U(L)11) gene of herpes simp lex virus type 1. This 96-amino-acid myristylated protein is thought to be involved in virion transport and envelopment at internal membrane sites. Wh en the first 100 amino acids of RSV Gag (including the M domain) were repla ced by the entire UL11 sequence, the chimeric protein localized at and budd ed into the Golgi apparatus father than being targeted to the plasma membra ne. Myristate was found to be required for this specific targeting, as were the first 49 amino acids of UL11, which contain an acidic fluster motif. I n addition to shedding new light on UL11, these experiments demonstrate tha t RSV Gag can be directed to internal cellular membranes and suggest that r egions outside of the M domain do not contain a dominant plasma membrane-ta rgeting motif.